The experiments described in this proposal are designed to gain insight into the nature of the control of gene expression in polyoma transformed cells. Since late gene expression is not normally observed in these cells, the cis and trans-acting factors that regulate transcription initiating from late promoter in integrated genomes will be determined by the analysis of the expression of chimeric plasmids that contain neomycin-coding sequences linked to this promoter. Cis elements influencing expression will be determined through the use of in vitro constructed mutants that contain deletions within the non-coding regulatory region. Molecular cloning and analysis of the structure of a polyoma insertion present within an anomalous cell line that expresses late mRNA's should also provide information on this subject. Chimeric plasmids containing the neomycin-resistance gene or the gene for chloramphenicol acetyl transferase will be used to determine if the polyoma large T antigen, when provided in trans, can influence the expression of these genes when they are linked to the late promoter in an integrated or extra-chromosomal state. Such a comparative analysis shousd provide information regarding the target and mechanism of this type of gene activation which recent evidence suggests is also operative in normal cells. Experiments designed to determine if the regulation of late gene expression in polyoma transformed cell lines occurs at a post-transcriptional level are also described. Finally, a temperature-inducible host-vector expression system is described. The characterization of this system will indicate the factors governing excision, gene amplification and expression in permissive mouse cells of foreign genes linked to the late promoter.